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80 × 40 Å with a narrow centrally attached stalk (15 Å wide and 100 Å long), which terminates into a
hydrophobic knob anchored in the viral envelope. The detergent-released spikes can be digested by
pronase to release the neuraminidase “heads,” which retain full antigenic and enzyme activity [40,41].
The molecule was found to be a tetramer of molecular weight 240,000, reducing to 200,000 when
treated with pronase [42]. A low resolution x-ray image of crystallized neuraminidase heads [43]
established that the enzyme had circular 4-fold symmetry.
III. Molecular Structure of Neuraminidase
Crystals of pronase-released heads of the N2 human strains of A/Tokyo/3/67 [44] and A/RI/5+/57 were
used for an x-ray structure determination. The x-ray 3-dimensional molecular structure of neuraminidase
heads was determined [45] for these two N2 subtypes by a novel technique of molecular electron density
averaging from two different crystal systems, using a combination of multiple isomorphous replacement
and noncrystallographic symmetry averaging. The structure of A/Tokyo/3/67 N2 has been refined [46]
to 2.2 Å as has the structures of two avian N9 subtypes [47–49]. Three influenza type B structures [50]
have also been determined and found to have an identical fold with 60 residues (including 16 conserved
cysteine residues) being invariant. Bacterial sialidases from salmonella [51] and cholera [52] have
homologous structures to influenza neuraminidase, but few of the residues are structurally invariant.
A. Structural Topology
The protein fold consists of a symmetric arrangement of six four-stranded antiparallel β sheets arrange
as blades of a propeller (Figure 3), the propeller axis being approximately parallel to but titled away
from the circular 4-fold axis of the tetramer. This tilt angle varies between the known subtypes. This
topology has now also been found in the seven β sheet propeller structure of bacterial methylamine
dehydrogenase [53] and galactose oxidase [54], and the eight β sheet propeller structure of methanol
dehydrogenase [55].
Each sheet of neuraminidase has a “W” topology (+1,+1,+1) [56] with four strands connected by reverse
turns (Figure 3). The first strand of each sheet enters from the top, approximately parallel to and near the
propeller axis; the fourth strand exits from the bottom, approximately perpendicular to the propeller axis.
Top and bottom surfaces of the head refer to the faces of the tetramer away and towards the viral
membrane respectively. Each sheet thus has a characteristic 90° right-hand twist between the inner- and
outermost strand. The six sheets and their connections to each other are topologically identical.
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